The long term objective of this proposal is to develop a fully integrated nanochannel chip and reader capable of single molecule mapping of linearized, long genomic material. The anticipated embodiment will permit direct visualization and analysis of megabase fragments of DNA extracted directly from a sample (possibly a single cell) with sub-kilobase resolution. Furthermore, the chip will accommodate massively parallel analyses of individual DNA molecules to permit standardized, high-throughput mapping of sequence motifs or polymorphic sites along the DNA. We propose a barcoding strategy (chip, reader and assay) for single DNA molecules. Our barcoding strategy is based on direct fluorescent imaging and localization of multiple sequence motifs or polymorphic sites along a linearized DNA molecule. Such capabilities will transform biological analyses, permitting highly sensitive detection of genetic loci for genome wide association studies, especially where haplotype information is required. In addition, mapping of pathogen genomes can also be performed. A critical consideration for the commercialization of this device is the consistent linearization and imaging of individual DNA molecules such that high resolution mapping of labeled sites can be performed. In light of this requirement, we propose a nanofluidic device in which individual DNA molecules are streamed and linearized in massively parallel nanoscale channels. The advantage of a nanochannel device is that DNA can be linearized in a standardized and repeatable manner due to the physical confinement of molecules within the channel. During linearization, the DNA is fluorescently imaged using a high resolution microscopy system thus permitting spatial mapping of site-specific fluorescent labels. Completion of this project will result in a chip, imaging system and assay for mapping sequence motifs and polymorphic sites along single molecules of DNA with 800 bp resolution. Such a device will transform and enhance our understanding of genetic diseases by providing crucial haplotype information. The device can be further extended to provide additional information regarding structural variations in the genome such as copy number variations and translocations.